The current tuberculosis vaccine, BCG, has questionable efficacy and undoubtedly new vaccines are needed. Recent studies have indicated that DNA vaccines may be promising as new mycobacterial vaccines candidates. We have generated 30 DNA vaccines expressing different mycobacterial antigens. All of these vaccines have been tested by intramuscular inoculation,either singly or in combination, for their capacity to induce protective, cytokine, and humoral immune responses in mice. Based on the results of these studies, we have concluded the following: 1. At least 10 tuberculosis DNA vaccines that we have tested induce protective immune responses in mice. However, the level of protection is less than the level of protection evoked by BCG. 2. Combinations of TB DNA vaccines can evoke a larger protective response than single vaccines. The level of protective immunity elicited by some combination TB DNA vaccines is similar to the BCG response. 3. Tuberculosis DNA vaccines encoding mycobacterial antigens fused to a TPA signal signal generally are more immunogenic than DNA vaccines encoding the native protein. 4. DNA vaccination induces a substantial splenic cytokine response and a lesser cytokine response in the lung. 5. Some DNA vaccines expressing mycobacterial antigens conjugated to ubiquitin evoke only cell-mediated and not humoral responses in mice. 6. Intranasal administration of TB DNA vaccines either in solution or encapsulated in liposomes does not induce an effective anti-tuberculosis protective response. We currently testing combinations of these TB DNA vaccines in long term studies to establish whether these preparations can induce extended protective immunity.